Characterization of fish skin mucus using mass spectrometry methods Investigation of biochemical differences in mucus of different fish species and sea lice

Abstract

The epidermal mucus layer of fish covers the entire exterior, including gills and fins. This barrier plays a crucial role in protecting and maintaining the skin heath, while it also reflects the fish’s welfare. The primary aims of this master’s thesis were to develop an in-lab methodology for analyzing fish skin mucus using Matrix Assisted Laser Desorption/ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) and to differentiate between species based on the protein content levels. It was also aimed to investigate distinctive variations in low molecular weight compounds within the mucus of salmonid species using Liquid Chromatography Mass Spectrometry (LC-MS). Using a purification method involving using zip-tip (C18), a form of SPE (solid-phase extraction) together with 60% acetonitrile (ACN) and 0.1% trifluoracetic (TFA), enhanced spectra on MALDI-TOF MS. In contrast, the methods involving untreated mucus, as well as mucus treated with dichloromethane, and with chloroform, yielded poorer spectra and was therefore not continued with. The superior purification method enabled the analysis and differentiation of Atlantic salmon, Atlantic cod, and coalfish based on the protein content levels in their skin mucus. Salmonid species could also be differentiated, although their varying spectra complicated the distinction of certain specimens. Skin mucus from salmonid species, along with whole lice, was analyzed using Liquid Chromatography Mass Spectrometry (LC-MS). The components of the mucus were depicted in a Principal Component Analysis (PCA) plot, which demonstrated that species tended to cluster more closely with its own type, rather than with others. The PCA-plot did not reveal any significant similarities between the sea lice, sea lice-infected specimens, and the non-infected specimens. The metabolites from the skin mucus of the salmonid species were compared through differential analysis. Although no notable difference between the sea liceinfected specimens and the non-infected specimens were observed in terms of metabolite composition, several other identified metabolites were found – which could in further research be interesting to proceed with. Ultimately, skin mucus remains a relatively unexplored topic, offering exciting opportunities in the future.